ORGANIC LETTERS
Daphlongeranines A and B, Two Novel Alkaloids Possessing Unprecedented Skeletons from Daphniphyllum longeracemosum
2007 Vol. 9, No. 13 2509-2512
Chun-Shun Li,†,‡ Ying-Tong Di,† Hong-Ping He,† Suo Gao,† Yue-Hu Wang,† Yang Lu,§ Jia-Liang Zhong,§ and Xiao-Jiang Hao*,† State Key Laboratory of Phytochemistry and Plant Resources in West China, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650204, Yunnan, PRC, Institute of Materia Medica, Chinese Academy of Medical Sciences, Beijing 100050, PRC, and Graduate School of Chinese Academy of Sciences, Beijing 100039, PRC
[email protected] Received April 22, 2007
ABSTRACT
Two novel alkaloids, daphlongeranines A (1) and B (2), with an unprecedented ring system, were isolated from the fruits of Daphniphyllum longeracemosum. Their unique structures and relative stereochemistries were established on the basis of spectroscopic and single-crystal diffraction analysis. The proposed biosynthetic pathway was also discussed. Compounds 1 and 2 showed weak inhibition on platelet aggregation.
Daphniphyllum alkaloids, elaborated by trees of the genus Daphniphyllum (Daphniphyllaceae), are a group of structurally diversified alkaloids with polycyclic skeletons,1-3 which have attracted great interest as challenging projects for total synthesis4 and biosynthetic research.5 In our continuing search for structurally unique and biogenetically interesting Daphniphyllum alkaloids,2,6 two * To whom correspondence should be addressed. Phone: (86) 8715223263. Fax: (86) 871-5219684. † Kunming Institute of Botany. ‡ Graduate School of Chinese Academy of Sciences. § Institute of Materia Medica. (1) For a review of Daphniphyllum alkaloids: Kobayashi, J.; Morita, H. In The Alkaloids; Cordell, G. A., Ed.; Academic Press: New York, 2003; Vol. 60, pp 165-205. (2) (a) Di, Y. T.; He, H. P.; Wang, Y. S.; Li, L. B.; Lu, Y.; Gong, J. B.; Fang, X.; Kong, N. C.; Li, S. L.; Zhu, H. J.; Hao, X. J. Org. Lett. 2007, 9, 1355-1358. (b) Li, C. S.; He, H. P.; Di, Y. T.; Wang, Y. H.; Mu, S. Z.; Li, S. L.; Gao, S.; Gao, Z. L.; Hao, X. J. Tetrahedron Lett. 2007, 48, 27372740. (c) Di, Y. T.; He, H. P.; Liu, H. Y.; Du, Z. Z.; Tian, J. M.; Yang, X. W.; Wang, Y. H.; Hao, X. J. Tetrahendron Lett. 2006, 47, 5329-5331. 10.1021/ol070942+ CCC: $37.00 Published on Web 06/01/2007
© 2007 American Chemical Society
novel alkaloids, daphlongeranines A (1) and B (2), with an unprecedented hepta- and hexacyclic ring system, respectively, were isolated. Herein, we present the isolation and structural elucidation of 1 and 2.
The air-dried and powdered fruits of D. longeracemosum Rosenth. were extracted with 95% EtOH, and the crude extract was adjusted to pH 2 with 2% HCl. After extracting with petroleum ether (PE) and CHCl3, the aqueous layer,
Figure 1. Key two-dimensional NMR correlations for daphlongeranines A (1) and B (2).
alkalinized to pH 10 with 3% NaOH, was then extracted with CHCl3. CHCl3-soluble materials were subjected to silica gel column chromatography (CHCl3/MeOH, 1:0 f 0:1), from which a fraction (CHCl3/MeOH, 20:1) was chromatographed over a series of silica gel columns to afford daphlongeranines A (1, 0.00035%) and B (2, 0.00025%). Daphlongeranine A (1)7 was isolated as a block crystal (PE/acetone 2:1). Its molecular formula was demonstrated as C24H31NO5 by HR-ESI-MS (m/z 414.2284, [M + H]+, calcd 414.2280), with ten degrees of unsaturation. The IR (3) (a) Saito, S.; Kutoba, T.; Fukushi, E.; Kawabata, J.; Zhang, H. P.; Kobayashi, J. Org. Lett. 2007, 9, 1207-1209. (b) Fan, C. Q.; Yin, S.; Xue, J. J.; Yue, J. M. Tetrahedron 2007, 63, 115-119. (c) Li, Z. Y.; Chen, P.; Xu, H. G.; Yang, Y. M.; Peng, S. Y.; Zhao, Z. Z.; Guo, Y. W. Org. Lett. 2007, 9, 477-480. (d) Zhang, W.; Guo, Y. W.; Krohn, K. Chem.-Eur. J. 2006, 12, 5122-5127. (4) (a) Wallace, G. A.; Heathcock, C. H. J. Org. Chem. 2001, 66, 450454. (b) Heathcock, C. H. Proc. Natl. Acad. Sci. U.S.A. 1996, 93, 1432314327. (c) Heathcock, C. H.; Joe, D. J. Org. Chem. 1995, 60, 1131-1142. (d) Heathcock, C. H.; Kath, J. C.; Ruggeri, R. B. J. Org. Chem. 1995, 60, 1120-1130. (5) (a) Niwa, H.; Hirata, Y.; Suzuki, K. T.; Yamamura, S. Tetrahedron Lett. 1973, 4, 2129-2132. (b) Suzuki, K. T.; Okuda, S.; Niwa, H.; Toda, M.; Hirata, Y.; Yamamura, S. Tetrahedron Lett. 1973, 14, 799-802. (6) (a) Mu, S. Z.; Yang, X. W.; Di, Y. T.; He, H. P.; Wang, Y.; Wang, Y. H.; Li, L.; Hao, X. J. Chem. BiodiVersity. 2007, 4, 129-138. (b) Li, L.; He, H. P.; Di, Y. T.; Gao, S.; Hao, X. J. Tetrahedron Lett. 2006, 47, 62596262. (c) Mu, S. Z.; Wang, Y.; He, H. P.; Yang, X. W.; Wang, Y. H.; Di, Y. T.; Lu, Y.; Chang, Y.; Hao, X. J. J. Nat. Prod. 2006, 69, 1065-1069. (d) Di, Y. T.; He, H. P.; Li, C. S.; Tian, J. M.; Mu, S. Z.; Li, S. L.; Gao, S.; Hao, X. J. J. Nat. Prod. 2006, 69, 1745-1748. (e) Li, L.; He, H. P.; Di, Y. T.; Tian, J. M.; Hao, X. J. HelV. Chim. Acta 2006, 89, 1457-1462. (7) Daphlongeranine A (1): light yellow block crystals (PE/acetone 2:1); mp 227-229 °C; [R]22D 142.7 (c 0.16, CHCl3); IR (KBr) νmax 3426, 2925, 1774, 1737, 1638, 1457, 1191, 755 cm-1; 1H and 13C NMR data, see Table 1; ESI-MS m/z 414 (M + 1)+; HR-ESI-MS m/z 414.2284 ([M + H]+; calcd for C24H31NO5, 414.2280). 2510
spectrum suggested the presence of a hydroxyl (3426 cm-1) and two carbonyl groups (1774 and 1737 cm-1). All 24 carbon signals observed in the 13C NMR and DEPT spectrum could be classified into one trisubstituted olefin, two carbonyls, four sp3 quaternaries, four sp3 methines, eleven sp3 methylenes together with one methyl. Among them, two methylenes (δC 64.2, δH 2.88 and 2.70; δC 45.8, δH 3.10 and 2.32) and one quaternary carbon (δC 72.7) were ascribed as attaching to a nitrogen atom, whereas one methylene (δC 64.4, δH 3.94 and 3.60) and two methines (δC 92.8, δH 4.26; δC 91.2, δH 4.58) were ascribed as bearing oxygen atoms. Besides the three degrees of unsaturation ascribed to two carbonyl groups and one olefin, seven degrees of unsaturation could only be assigned to the presence of a heptacyclic skeleton. The structure of 1 was elucidated to be composed of two moieties (Figure 1) by analyses of 2D NMR (1H-1H COSY, HSQC, and HMBC). The 1H-1H COSY spectrum revealed connectivity of two structural fragments: a (C-6 to C-7 and C-12, C-10 to C-12 and C-17, C-15 to C-17), and b (C-13 to C-14) drawn with bold bonds in Figure 1. Furthermore, HMBC correlations proved the presence of the right moiety, as shown in Figure 1 (in red), containing two fused ring systems (ring A and B) with a double bond between C-9 and C-15, a propionate at C-8, an oxomethylene at C-5, and a methylene at C-6, which were the common structural units in yuzurimine- and daphnilactone-type alkaloids.1 In the left moiety, HMBC correlations of H-7R (δH 2.32) and H2-19 (δH 2.88 and 2.70) with C-3 (δC 72.7) and of H2-7 with C-19 (δC 64.2) indicated that C-3, C-7, and C-19 Org. Lett., Vol. 9, No. 13, 2007
connected to each other through the nitrogen atom. Correlation signals of H2-4 to C-3 (2J) and C-5 (2J), as well as of H2-7 to C-5 (3J), demonstrated the existence of a sixmembered heterocyclic ring C (C-3, C-4, C-5, C-6, C-7, and nitrogen). Meanwhile, the formation of ring D was similarly verified by correlations of H-1 to C-8 and of H2-4 to C-1 and C-8, respectively. HMBC cross-peaks of H-2 (δH 4.26) with C-3, C-18, and C-19, respectively, along with the correlations of H2-19 to C-18 indicated the existence of ring E (C-2, C-3, nitrogen, C-19, and C-18). The presence of the five-membered lactone ring G (C-2, C-18, C-23, C-24, and oxygen) and assignment of the methyl (C-20, δC 24.7) at C-18 were achieved by the 3J correlations of H-2 to C-23 and C-24 and the 2J correlations of H2-23 to C-18 and C-24, as well as of H3-20 to C-2 (3J). However, the correlation between H-1 and C-22 could not be observed in the HMBC spectrum; therefore, the formation of δ-lactone ring F was eventually resolved by the X-ray experiment. Thus, the gross structure of daphlongeranine A (1), possessing a unique heptacyclic ring system with two lactones, was established as shown in Figure 1. The relative stereochemistry of 1 was elucidated by analysis of the ROESY spectrum as shown in Figure 1 and was further confirmed by an X-ray crystallographic study.8 In the crystal structure (Figure 2), H-2, H-6, and H-10 took
E, and G took an envelope conformation; the rings B and C took a chair conformation; and the ring F took a twist-boat conformation. Daphlongeranine B (2)9 was obtained as needle crystals (PE/acetone 2:1). Its molecular formula, C22H29NO4, was established by HR-ESI-MS (m/z 372.2174, [M + H]+, calcd 372.2174), corresponding to nine degrees of unsaturation. The IR spectrum indicated the absorption of carbonyl (1755 cm-1) and hydroxyl (3439 cm-1) groups. Extensive comparison of 1H NMR, 13C NMR, and DEPT spectra of compound 2 with 1 (see Table 1) suggested that
Table 1. 1H (400 MHz) and 13C (100 MHz) NMR Data of Daphlongeranines A (1)a and B (2)b no.
δHa
δCa
δHb
δCb
1 2 3 4a 4b 5 6 7R 7β 8 9 10 11 12
4.58 (1H, d, 1.6) 4.26 (1H, s) --1.66 (1H, dd, 2.0, 12.0) 1.52 (1H, d, 12.0) --2.10-2.04 (1H, m) 2.32 (1H, m) 3.10 (1H, t, 14.0) ----2.56-2.52 (1H, m) 1.52-1.37 (2H, m) 1.30 (2H, m)
91.2 92.8 72.7 32.5
85.0 211.4 72.7 39.3
13a 13b 14a 14b 15 16R 16β 17R 17β 18 19R 19β 20 21a 21b 22 23R 23β 24
1.90-1.81 (1H, m) 1.30-1.25 (1H, m) 2.35-2.28 (1H, m) 2.19-2.15 (1H, m) 5.38 (1H, s) 2.10-2.04 (1H, m) 2.33-2.29 (1H, m) 1.52-1.50 (1H, m) 1.89-1.81 (1H, m) --2.70 (1H, d, 9.2) 2.88 (1H, d, 9.2) 1.18 (3H, s) 3.94 (1H, d, 10.8) 3.60 (1H, d, 10.8) --2.55 (1H, d, 18.0) 2.45 (1H, d, 18.0) ---
4.67 (1H, d, 9.2) ----2.36 (1H, m) 1.43 (1H, m) --2.25 (1H, m) 2.58 (1H, m) 2.73 (1H, dd, 4.8, 11.2) ----2.57 (1H, m) 1.65-1.49 (2H, m) 1.92 (1H, m) 1.52 (1H, m) 2.00 (1H, m) 1.34 (1H, m) 2.42-2.27 (2H, m)
126.9 29.3
173.7 43.9
5.29 (1H, s) 2.17 (1H, m) 2.43 (1H, m) 1.61 (1H, dd, 6.8, 12.4) 1.96 (1H, m) 2.51 (1H, m) 2.92 (1H, t, 9.0) 2.78 (1H, dd, 1.4, 9.0) 1.16 (1H, d, 7.6) 4.07 (1H, d, 11.0) 3.77 (1H, d, 11.0) -----
177.5
---
a
52.3 35.1 45.8 49.2 151.1 44.9 30.2 24.8 25.8 26.4 126.0 29.1 34.8 42.2 64.2 24.7 64.4
52.2 40.1 50.6 50.0 151.1 44.7 31.5 25.8 24.9 25.9
34.8 39.8 54.4 16.1 64.8 171.6 -----
b
NMR data for 1 (in CDCl3:CD3OD, 9:1). NMR data for 2 (in CDCl3).
Figure 2. X-ray crystal structure of 1 (ORTEP drawing).
the β configuration, and H-1 was R facial. The rings A, D, (8) Crystallographic data of 1: C24H31NO5, M ) 413.51, monoclinic, space group P21, a ) 13.576(1) Å, b ) 8.049(2) Å, c ) 10.772(1) Å, β ) 109.893(4)°, V ) 1106.9(2) Å3, Z ) 2, crystal dimensions 0.25 × 0.40 × 0.60 mm3 were used for measurements on a MAC DIP-2030K diffractometer with a graphite monochromator (ω scan, 2θ max ) 50.0°), Mo KR radiation. The total number of independent reflections measured was 2124, of which 2093 were observed (|F|2 G 2σ|F|2). Final indices: Rf ) 0.0381, wR2 ) 0.1026 (w ) 1/σ|F|2), S ) 1.050. The crystal structure (1) was solved by the direct method SHELXS-97, expanded using geometrical calculations and difference Fourier techniques, and refined by least-squares calculations. Crystallographic data for the structure of 1 have been deposited in the Cambridge Crystallographic Date Centre (deposition number: CCDC 634546). Copies of this data can be obtained free of charge via http:// www.ccdc.cam.ac.uk/deposit (or from Cambridge Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ, U.K., facsimile: (44) 01223 336033, e-mail:
[email protected]).
Org. Lett., Vol. 9, No. 13, 2007
2 had a skeleton partially similar to 1. The presence of rings A-D and F in alkaloid 2 was further demonstrated by 2D NMR (1H-1H COSY, TOCSY, HSQC, and HMBC) data similar to compound 1, as shown in Figure 1. The 1H-1H COSY and TOCSY spectra (Figure 1) of 2 implied the presence of unit c (C-18 to C-19), and HMBC correlations of H3-20 to C-2 (3J), C-18 (2J) and C-19 (2J), H-18 and H-19R to C-2 established the formation of ring E with C-20 at C-18. Thus, the gross structure of 2 with a skeleton similar to 1 was assigned as shown in Figure 1. (9) Daphlongeranine B (2): colorless needle crystals (PE/acetone 2:1); mp 241-243 °C; [R]17D 47.2 (c 0.41); IR (KBr) νmax 3439, 2931, 1755, 1637, 1455, 1052, 756 cm-1; 1H and 13C NMR data, see Table 1; ESI-MS m/z 372 (M + 1)+; HR-ESI-MS m/z 372.2174 ([M + H]+; calcd for C22H29NO4, 372.2174). 2511
Scheme 1.
Plausible Biosynthetic Pathway of 1 and 2
The relative configuration of compoud 2, as shown in Figure 1, was deduced from its ROESY spectrum and the comparison of NMR data with 1. Correlations of H2-21/H213, H-4a, H-6, and H-10, and H-15/H-1 indicated that alkaloid 2 had the same configuration as 1. Meanwhile, the NOE correlation of H-4a/H-20 implied that Me-18 (C-20) was in β configuration. Due to the structural similarity between compounds 1 and 2, the latter should be the precursor of the former. Therefore, a plausible biogenetic pathway for them is proposed as shown in Scheme 1. The biosynthetic precursor of both 1 and 2 could be admitted to caldaphnidine C,10 which should produce a key intermediate C through a pinacol rearrangement.2c The oxidation of intermediate C at C-3 followed by further rearrangement might produce daphlongeranine B (2) through the formation of a C-3-N bond11 and the following esterification. After an aldol-type condensation reaction with pyruvic acid and subsequent dehydration and decarboxyla(10) Zhang, Z. J.; Zhang, C. R.; Yue, J. M. Tetrahedron 2005, 61, 11038-11045. (11) Saito, S.; Kubita, T.; Fukushi, E.; Kawabata, J.; Zhang, H. P.; Kobayashi, J. Tetrahedron Lett. 2007, 48, 1587-1589.
2512
tion, daphlongeranine B (2) will eventually convert to daphlongeranine A (1). Compounds 1 and 2 showed weak inhibition on vitro platelet aggregation induced by ADP, PAF, and AA. At 240 µM concentration, the inhibition of aggregation for 1 was 38.4 ( 17.2%, 37.5 ( 14.7%, and 19.8 ( 7.0% and for 2 was 38.7 ( 10.5%, 28.3 ( 9.4%, and 24.0 ( 11.2%, respectively. Acknowledgment. The authors thank Prof. Xun Gong, Kunming Institute of Botany, Chinese Academy of Sciences (CAS), for the identification of the plant material. This work was supported by the National Science Foundation (No. 20672120) of PRC. Supporting Information Available: Experimental procedures, 1D and 2D NMR spectra for compounds 1 and 2, and X-ray crystallographic data of compound 1 (PDF and CIF). This material is available free of charge via the Internet at http://pubs.acs.org. OL070942+
Org. Lett., Vol. 9, No. 13, 2007
